RESEARCH ACCOMPLISHMENTS A. CHARACTIZATION OF THE NOVEL INTERFERON-MEDIATED AUTOINFLAMMATORY DISEASE CAUSED BY GAIN OF FUNCITON MUTATIONS IN STING, SAVI. We recently identified that gain of function mutations in TMEM173, the gene that encodes a key adaptor molecule in the interferon pathway, STING, causes a novel autoinflammatory disease that we named STING-associated vasculopathy with onset in infancy (SAVI). These patients have constitutive activation of the interferon-beta pathway. Patients present with vasculitis and a severity spectrum of interstitial lung disease, and several patients have developed pulmonary hypertension. STING is expressed in endothelial cells, and activation of the STING pathways in these cells results in endothelial activation and apoptosis, a feature that may explain the predominant manifestation as peripheral vasculitis and vasculopathy. However the severity of interstitial lung disease is variable in patients with SAVI ranging from being absent to severe. We observed that activated/phosphorylated STAT1 in SAVI patients lymphocytes and the exaggerated responses to STING agonists including cGAMP in patient fibroblasts are reduced by treatment with JAK inhibitors, we are treating SAVI patients in a compassionate use study with the JAK inhibitor baricitinib and evaluate the effect of treatment on the clinical manifestations including vasculitis, interstitial lung disease and pulmonary hypertension. We are characterizing genetic variants that modify the severity of the IFN dysregulation and are hypothesizing that these variants may determine the severity of the lung disease in SAVI patients. B. GAIN OF FUNCTION MUTATION IN NLRC4 CAUSE AN AUTOINFLAMMATORY SYNDROME WITH RECURRENT FEVER FLARES AND MACROPHAGE ACTIVATION (NLRC4-MAS) The identification of a de novo mutation T337S in the intracellular innate immune sensor, NLRC4 identified a monogenic defect that can lead to the development of macrophage activation syndrome. Functional analyses pf the mutation demonstrate spontaneous inflammasome formation and production of the inflammasome-dependent cytokines IL-1beta and IL-18. In contrast to NOMID patients with an inflammasome defect in NLRP3, NLRC4-MAS patients have very high elevation of IL-18 produced in macrophages, thus providing us with a disease model to study macrophage activation. High IL-18 levels are also seen in patients with other inflammatory conditions that are prone to the development of macrophage activation syndrome i.e Stills disease and our data suggest a critical role for IL-18 in the development of macrophage activation syndrome. This project is being developed in collaboration with the Metzger Scholar, Dr. Canna. C. NOVEL INSIGHTS INTO THE INTERFERON MEDIATED DISEASE CANDLE. The majority of patients with CANDLE (Chronic atypical neutrophilic dermatosis with lipodystrophy, and elevated temperature) have mutations in the proteasome subunit PSMB8 which led to the unifying concept that a group of rare diseases referred to as JMP (joint contractures, muscle atrophy and panniculitis-induced lipodystrophy) syndrome, and Nakajo-Nishimura syndrome (NNS) form one disease spectrum of proteasome-associated autoinflammatory syndromes. We have recently discovered that CANDLE patients who were negative for PSMB8 mutations have mutations in other proteasome components that lead to proteasome dysfunction indicating that CANDLE can be caused by digenic and monogenic inheritance. All patients expressed high IP-10 (Interferon gamma-induced protein 10), MCP-1, and other chemokines and cytokines associated with interferon induced diseases and a strong interferon (IFN) response signature (IRS) confirming that IFN dysregulation is linked to global proteasome dysfunction not related to mutations in one component. An ongoing compassionate use study with the JAK inhibitor, baricitinib (Eli Lilly) that can inhibit interferon signaling, is recruiting patients with CANDLE and SAVI and other interferonopathies who are unresponsive to IL-1 blocking agents. We are studying the inflammatory phenotype and metabolic changes in CANDLE, including hyperlipidemias, metabolic syndrome, myositis and primary pulmonary hypertension, a serious disease manifestation we recently identified in CANDLE patients. D. STUDIES ON THE IL-1-MEDIATED DISEASES, NOMID AND DIRA: 1. FDA submission of our long-term outcome data using the short-acting IL-1 inhibitor anakinra (SOBI) in patients with neonatal-onset multisystem inflammatory disease (NOMID) led to FDA approval of anakinra for the treatment of NOMID in December 2012. 2. We previously found that IL-1 blocking treatment adjustments need to be made to control central nervous system (CNS) inflammation in NOMID patients. We thus evaluated biomarkers in the cerebrospinal fluid (CSF) that correlate with CNS inflammation and suggest a contribution on non hematopoietic cells to the inflammatory response in the CNS. We validated these markers in patients treated with a short-acting and long-acting IL-1 inhibitor. 3. Dr. Montealegre in our group conducts a study with the long-acting IL-1 blocking agent rilonacept in patients with DIRA. Preliminary data suggest good inflammatory control and prevention of progression of organ damage in six patients enrolled. E. ONGING STUDY OF PATIENTS WITH UNDIFFERENTIATED AUTOINFLAMMTORY DISEASES We evaluate and treat patients with severe inflammatory diseases that present early in infancy. In addition to a detailed immune evaluation that includes gene expression studies, patients and their parents if indicated undergo genetic analyses including evaluation through next generation sequencing, including whole exome sequencing (WES). We have identified likely disease causing mutations that are being evaluated and we attempt to identify molecular targets that allow us to better treat patients with AIDs. CONCLUSIONS AND SIGNIFICANCE We developed an integrative approach using immune evaluation, whole exome sequencing and targeted therapeutics to assess patients with autoinflammatory disorders, with the goal to better diagnose, understand disease pathogenesis and identify novel targets for therapy. Our studies have resulted in the discovery of the genetic causes of novel diseases that provided insights into the disease pathogenesis and targets for novel treatment interventions. 1. Two novel recently identified autoinflammatory diseases, SAVI and NLRC4-MAS shed light on basic disease mechanisms that cause autoinflammatory disease phenotypes. 2. Our pathogenesis data in CANDLE and our findings that gain-of-function mutations in STING, a gatekeeper for IFN beta transcription cause SAVI, suggest a causative role of IFN signaling in a new group of autoinflammatory diseases and suggest novel targets for treatment. 3. Our studies in CANDL and SAVI provide a rational for a compassionate use study with the JAK1/2 inhibitor baricitinib that inhibits IFN signaling. We are assessing the clinical efficacy and safety in patients with CANDLE and SAVI and in patients with clinical and laboratory evidence of IFN mediated disease. 4. The identification mutations in NLRC4 causing macrophage activation, lead to the exploration of the role of IL-18 in macrophage activation and suggest IL-18 as a potential target for treatment. 5. IL-1-blocking therapies are FDA-approved for the use in CAPS including NOMID based on work performed under this study. Our biomarker analyses aim to define markers that help us determine appropriate control of systemic and organ inflammation in NOMID and to identify potential differences in the effectiveness in controlling organ inflammation that may exist between different IL-1 blocking agents. 6. A clinical study in patients with DIRA using the long-acting IL-1 inhibitor rilonacept (Regeneron) is being completed.